Neuroprotection by post-stroke administration of an oral formulation of angiotensin-(1-7) in ischaemic stroke.

NEW FINDINGS: What is the central question of this study? Angiotensin-(1-7) decreases cerebral infarct volume and improves neurological function when delivered centrally before and during ischaemic stroke. Here, we assessed the neuroprotective effects of angiotensin-(1-7) when delivered orally post-stroke. What is the main finding and its importance? We show that oral delivery of angiotensin-(1-7) attenuates cerebral damage induced by middle cerebral artery occlusion in rats, without affecting blood pressure or cerebral blood flow. Importantly, these treatments begin post-stroke at times coincident with the treatment window for tissue plasminogen activator, providing supporting evidence for clinical translation of this new therapeutic strategy. ABSTRACT: As a target for stroke therapies, the angiotensin-converting enzyme 2-angiotensin-(1-7)-Mas [ACE2/Ang-(1-7)/Mas] axis of the renin-angiotensin system can be activated chronically to induce neuroprotective effects, in opposition to the deleterious effects of angiotensin II via its type 1 receptor. However, more clinically relevant treatment protocols with Ang-(1-7) that involve its systemic administration beginning after the onset of ischaemia have not been tested. In this study, we tested systemic post-stroke treatments using a molecule where Ang-(1-7) is included within hydroxypropyl-ß-cyclodextrin [HPßCD-Ang-(1-7)] as an orally bioavailable treatment. In three separate protocols, HPßCD-Ang-(1-7) was administered orally to Sprague-Dawley rats after induction of ischaemic stroke by endothelin-1-induced middle cerebral artery occlusion: (i) to assess its effects on cerebral damage and behavioural deficits; (ii) to determine its effects on cardiovascular parameters; and (iii) to determine whether it altered cerebral blood flow. The results indicate that post-stroke oral administration of HPßCD-Ang-(1-7) resulted in 25% reductions in cerebral infarct volumes and improvement in neurological functions (P < 0.05), without inducing any alterations in blood pressure, heart rate or cerebral blood flow. In conclusion, Ang-(1-7) treatment using an oral formulation after the onset of ischaemia induces significant neuroprotection in stroke and might represent a viable approach for taking advantage of the protective ACE2/Ang-(1-7)/Mas axis in this disease.

Role of neurons and glia in the CNS actions of the renin-angiotensin system in cardiovascular control.

Despite tremendous research efforts, hypertension remains an epidemic health concern, leading often to the development of cardiovascular disease. It is well established that in many instances, the brain plays an important role in the onset and progression of hypertension via activation of the sympathetic nervous system. Further, the activity of the renin-angiotensin system (RAS) and of glial cell-mediated proinflammatory processes have independently been linked to this neural control and are, as a consequence, both attractive targets for the development of antihypertensive therapeutics. Although it is clear that the predominant effector peptide of the RAS, ANG II, activates its type-1 receptor on neurons to mediate some of its hypertensive actions, additional nuances of this brain RAS control of blood pressure are constantly being uncovered. One of these complexities is that the RAS is now thought to impact cardiovascular control, in part, via facilitating a glial cell-dependent proinflammatory milieu within cardiovascular control centers. Another complexity is that the newly characterized antihypertensive limbs of the RAS are now recognized to, in many cases, antagonize the prohypertensive ANG II type 1 receptor (AT1R)-mediated effects. That being said, the mechanism by which the RAS, glia, and neurons interact to regulate blood pressure is an active area of ongoing research. Here, we review the current understanding of these interactions and present a hypothetical model of how these exchanges may ultimately regulate cardiovascular function.

Brain angiotensin type-1 and type-2 receptors: cellular locations under normal and hypertensive conditions.

Brain angiotensin-II (Ang-II) type-1 receptors (AT1Rs), which exert profound effects on normal cardiovascular, fluid, and metabolic homeostasis, are overactivated in and contribute to chronic sympathoexcitation and hypertension. Accumulating evidence indicates that the activation of Ang-II type-2 receptors (AT2Rs) in the brain exerts effects that are opposite to those of AT1Rs, lowering blood pressure, and reducing hypertension. Thus, it would be interesting to understand the relative cellular localization of AT1R and AT2R in the brain under normal conditions and whether this localization changes during hypertension. Here, we developed a novel AT1aR-tdTomato reporter mouse strain in which the location of brain AT1aR was largely consistent with that determined in the previous studies. This AT1aR-tdTomato reporter mouse strain was crossed with our previously described AT2R-eGFP reporter mouse strain to yield a novel dual AT1aR/AT2R reporter mouse strain, which allowed us to determine that AT1aR and AT2R are primarily localized to different populations of neurons in brain regions controlling cardiovascular, fluid, and metabolic homeostasis. Using the individual AT1aR-tdTomato reporter mice, we also demonstrated that during hypertension induced by the administration of deoxycorticosterone acetate-salt, there was no shift in the expression of AT1aR from neurons to microglia or astrocytes in the paraventricular nucleus, a brain area important for sympathetic regulation. Using AT2R-eGFP reporter mice under similar hypertensive conditions, we demonstrated that the same was true of AT2R expression in the nucleus of the solitary tract (NTS), an area critical for baroreflex control. Collectively, these findings provided a novel means to assess the colocalization of AT1R and AT2R in the brain and a novel view of their cellular localization in hypertension.

Butyrate regulates inflammatory cytokine expression without affecting oxidative respiration in primary astrocytes from spontaneously hypertensive rats.

Neurons and glia exhibit metabolic imbalances in hypertensive animal models, and loss of metabolic homeostasis can lead to neuroinflammation and oxidative stress. The objective of this study was to determine the effects of the microbial metabolite butyrate on mitochondrial bioenergetics and inflammatory markers in mixed brainstem and hypothalamic primary cultures of astrocytes between normotensive (Sprague-Dawley, S-D) and spontaneously hypertensive (SHR) rats. Bioenergetics of mitochondria in astrocytes from normotensive S-D rats were modified with butyrate, but this was not the case in astrocytes derived from SHR, suggesting aberrant mitochondrial function. Transcripts related to oxidative stress, butyrate transporters, butyrate metabolism, and neuroinflammation were quantified in astrocyte cultures treated with butyrate at 0, 200, 600, and 1000 µmol/L. Butyrate decreased catalase and monocarboxylate transporter 1 mRNA in astrocytes of S-D rats but not in the SHR. Moreover, while butyrate did not directly regulate the expression of 3-hydroxybutyrate dehydrogenase 1 and 2 in astrocytes of either strain, the expression levels for these transcripts in untreated cultures were lower in the SHR compared to S-D. We observed higher levels of specific inflammatory cytokines in astrocytes of SHR, and treatment with butyrate decreased expression of Ccl2 and Tlr4 in SHR astrocytes only. Conversely, butyrate treatment increased expression of tumor necrosis factor in astrocytes from SHR but not from the S-D rats. This study improves our understanding of the role of microbial metabolites in regulating astrocyte function, and provides support that butyrate differentially regulates both the bioenergetics and transcripts related to neuroinflammation in astrocytes from SHR versus S-D rats.

Gut dysbiosis is linked to hypertension.

Emerging evidence suggests that gut microbiota is critical in the maintenance of physiological homeostasis. This study was designed to test the hypothesis that dysbiosis in gut microbiota is associated with hypertension because genetic, environmental, and dietary factors profoundly influence both gut microbiota and blood pressure. Bacterial DNA from fecal samples of 2 rat models of hypertension and a small cohort of patients was used for bacterial genomic analysis. We observed a significant decrease in microbial richness, diversity, and evenness in the spontaneously hypertensive rat, in addition to an increased Firmicutes/Bacteroidetes ratio. These changes were accompanied by decreases in acetate- and butyrate-producing bacteria. In addition, the microbiota of a small cohort of human hypertensive patients was found to follow a similar dysbiotic pattern, as it was less rich and diverse than that of control subjects. Similar changes in gut microbiota were observed in the chronic angiotensin II infusion rat model, most notably decreased microbial richness and an increased Firmicutes/Bacteroidetes ratio. In this model, we evaluated the efficacy of oral minocycline in restoring gut microbiota. In addition to attenuating high blood pressure, minocycline was able to rebalance the dysbiotic hypertension gut microbiota by reducing the Firmicutes/Bacteroidetes ratio. These observations demonstrate that high blood pressure is associated with gut microbiota dysbiosis, both in animal and human hypertension. They suggest that dietary intervention to correct gut microbiota could be an innovative nutritional therapeutic strategy for hypertension.

The angiotensin type 2 receptor agonist Compound 21 elicits cerebroprotection in endothelin-1 induced ischemic stroke.

Evidence indicates that angiotensin II type 2 receptors (AT2R) exert cerebroprotective actions during stroke. A selective non-peptide AT2R agonist, Compound 21 (C21), has been shown to exert beneficial effects in models of cardiac and renal disease, as well as hemorrhagic stroke. Here, we hypothesize that C21 may exert beneficial effects against cerebral damage and neurological deficits produced by ischemic stroke. We determined the effects of central and peripheral administration of C21 on the cerebral damage and neurological deficits in rats elicited by endothelin-1 induced middle cerebral artery occlusion (MCAO), a model of cerebral ischemia. Rats infused centrally (intracerebroventricular) with C21 before endothelin-1 induced MCAO exhibited significant reductions in cerebral infarct size and the neurological deficits produced by cerebral ischemia. Similar cerebroprotection was obtained in rats injected systemically (intraperitoneal) with C21 either before or after endothelin-1 induced MCAO. The protective effects of C21 were reversed by central administration of an AT2R inhibitor, PD123319. While C21 did not alter cerebral blood flow at the doses used here, peripheral post-stroke administration of this agent significantly attenuated the MCAO-induced increases in inducible nitric oxide synthase, chemokine (C-C) motif ligand 2 and C-C chemokine receptor type 2 mRNAs in the cerebral cortex, indicating that the cerebroprotective action is associated with an anti-inflammatory effect. These results strengthen the view that AT2R agonists may have potential therapeutic value in ischemic stroke, and provide the first evidence of cerebroprotection induced by systemic post stroke administration of a selective AT2R agonist.